title: A rapid and cost-effective method of producing recombinant proBNP and NT-proBNP variants in Escherichia coli for immunoassay of heart failure.
abstract: The measurements of plasma natriuretic peptides (NT-proBNP, proBNP and BNP) are used to diagnose heart failure but these are expensive to produce. We describe a rapid, cheap and facile production of proteins for immunoassays of heart failure. DNA encoding N-terminally His-tagged NT-proBNP and proBNP were cloned into the pJexpress404 vector. ProBNP and NT-proBNP peptides were expressed in Escherichia coli, purified and refolded in vitro. The analytical performance of these peptides were comparable with commercial analytes (NT-proBNP EC50 for the recombinant is 2.6 ng/ml and for the commercial material is 5.3 ng/ml) and the EC50 for recombinant and commercial proBNP, are 3.6 and 5.7 ng/ml respectively). Total yield of purified refolded NT-proBNP peptide was 1.75 mg/l and proBNP was 0.088 mg/l. This approach may also be useful in expressing other protein analytes for immunoassay applications.
  
  
  OBJECTIVE
  To develop a cost effective protein expression method in E. coli to obtain high yields of NT-proBNP (1.75 mg/l) and proBNP (0.088 mg/l) peptides for immunoassay use.
authors.affiliation: Saliva Translational Research Group, The Translational Research Institute, The University of Queensland Diamantina Institute, Level 6, TRI, 37 Kent St, Woolloongabba, QLD, 4102, Australia.
authors.forename: Muhammad Tarmizi
authors.initials: MT
authors.lastname: Soleh
authors.forename: Jared Yong Yang
authors.initials: JY
authors.lastname: Foo
authors.forename: Ulla-Maja
authors.initials: UM
authors.lastname: Bailey
authors.forename: Nikki Yi
authors.initials: NY
authors.lastname: Tan
authors.forename: Yunxia
authors.initials: Y
authors.lastname: Wan
authors.forename: Justin
authors.initials: J
authors.lastname: Cooper-White
authors.forename: Benjamin Luke
authors.initials: BL
authors.lastname: Schulz
authors.forename: Chamindie
authors.initials: C
authors.lastname: Punyadeera
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D015415:Biomarkers; D004797:Enzyme-Linked Immunosorbent Assay / Q000295:instrumentation*; D004926:Escherichia coli / Q000235:genetics*; D006333:Heart Failure; D006801:Humans; D008969:Molecular Sequence Data; D020097:Natriuretic Peptide, Brain / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015203:Reproducibility of Results
chemical_list: D015415:Biomarkers; D010446:Peptide Fragments; D011994:Recombinant Proteins; C109794:pro-brain natriuretic peptide (1-76); D020097:Natriuretic Peptide, Brain